This study of 17-beta-estradiol (E2) as a repressor of vein graft intimal hyperplasia (IH) is part of the long-term goal of finding clinical treatments to suppress the expression of genes associated with fibroproliferative disease. Early and persistent infiltration of macrophages into the vein wall occurs coincident with the upregulation of monocyte chemoattractant protein-1 (MCP-1). Many proteins associated with inflammation, including MCP-1, are under the control of nuclear factor kappaB (NF-kB). We have shown that E2 treatment reduces vein graft IH and hypothesize that E2 is decreasing the gene expression of inflammatory mediators and that this effect is secondary to inhibition of NF-kB. Specific Aim I: Evaluate E2 for a dose dependent effect on IH and macrophage accumulation in a rat femoral artery interposition vein graft. Five groups of male rats will have pellets implanted s.c. to achieve continuous E2 serum levels from 0 to 400 pg/ml. Changes in vein wall areas and inflammatory infiltrates will be assessed histologically and immunohistochemically respectively. Optimum E2 dose will be chosen for use in Specific Aims II and III. Specific Aim II: Measure the influence of the efficacious E2 treatment on the message and protein levels of MCP-1, TNF-alpha, and iNOS. These genes are known to be targets of NF-kB. Group 1 receives no E2; Group 2 receives E2 for 12 weeks, and Group 3 will have their E2 pellet excised at week 6 of 12 weeks. Changes in mRNA will be calculated with quantitative PCR and confirmed using RNAse protection assays. Blood serum E2 levels will be correlated with graft morphology and inflammatory mRNA and protein levels over time by analysis of variance. Specific Aim III: Investigate the estrogen receptor (ER)/NF-kB interaction and compare this interaction to the interaction of ER to a putative estrogen response element and to the interaction of ER with other transcription factors known to be influenced by ER. To determine if E2 blocks NF-kB binding to DNA, we will perform electrophoretic mobility shift assays. To screen for system components, antibodies to known cofactors of NF-kB will be evaluated for their ability to super shift the NF-kB complex with and without the presence of E2. Coimmunoprecipitation experiments will screen for proteins associated with ER or NF-kB, which may lead to an understanding of the molecular mechanism of the interaction between E2 and inflammation. Defining these mechanisms/mediators will provide information for devising new therapeutic strategies for control of IH.